Two types of naturally occurring polyunsaturated fatty acids (PUFA) can be distinguished, based on the relative position of double bonds: i.e. (i) fatty acids with isolated double bonds, such as linoleic acid (C18:2), and (ii) fatty acids having conjugated double bonds. Linoleic acid is a major component in vegetable oils, with soybean and safflower oil typically containing linoleic acid levels up to 53.7% and 77.7%, respectively. Fatty acids with conjugated double bonds occur relatively rarely in natural fats. However, conjugated fatty acids have gained increasing importance in recent years because of their nutritional and technological properties.
From a technological point of view, conjugated fatty acids find use as drying oils in paints, varnishes and plastics. Drying oils polymerize or “dry” after they have been applied to a surface to form tough, adherent and abrasion resistant films. As conjugated double bonds are more reactive than unconjugated ones, CLAs are also used as co-monomers in the production of bio-plastics via cationic and free radical copolymerization. Tung oil is an example of a naturally occurring oil containing significant levels of conjugated fatty acids. Because tung oil is expensive for many industrial applications, research was conducted in order to find substitutes.
In the particular case of conjugated linoleic acid (CLA), i.e. the positional and geometric conjugated dienoic isomers of linoleic acid (C18:2, which can have either the cis or trans configuration), the cis-9,trans-11 isomer of CLA (c9,t11-CLA, rumenic acid) is by far the predominant form in foods (as much as 90% of the total CLA content), especially in milk and tissue fat of ruminants. Synthetic CLA mixtures consist mostly of c9,t11- and t10,c12-CLA in almost equal amounts, with traces of t9,t11- and t10,t12-CLAs.
A variety of positive health effects have been attributed to CLAs. They are claimed to be anticarcinogenic, antidiabetic, antioxidative and antiarteriosclerotic. They decrease fat and increase muscle content in the body, reduce inflammation, show a beneficial effect on bone formation, enhance immune functions and reduce asthma (Bhattacharya et al, 2006). Recent studies conducted with enriched preparations in either c9,t11- or either t10,c12-CLA show that the two isomers assess different biological activity. Several reviews summarize the health effects of CLA mixtures and the purified form of c9,t11- and t10c12-CLA isomers (Bhattacharya et al., 2006; Pariza et al., 2001). Next to the c9,t11- and the t10,c12-CLA isomer, the t9,t11-CLA also exhibit beneficial health effects, which are superior compared with the more abundant c9,t11- and t10,c12-CLAs (Coakley et al., 2006; Ecker et al., 2009; Lee & Vanden Heuvel, 2010).
Because of their technological and nutritional applications, several isomerisation reactions to convert fatty acids having isolated double bonds into fatty acids having conjugated double bonds have been developed.
Conjugated PUFA Production Via Alkaline Isomerisation
In the alkali isomerization process, PUFA, such as linoleic acid, or an oil rich in PUFA or linoleic acid, like safflower oil, is treated at high temperature (200-250° C.) under alkaline conditions and an inert atmosphere (N2). In the current commercial processes strong bases, like NaOH or KOH, dissolved in water, are used. During the alkaline treatment isomerization as well as saponification takes place. As a consequence, the triglyceride structure is broken and glycerol and soaps are formed. Afterwards the aqueous phase, containing glycerol and the homogeneous base, is separated and the soaps are treated with an acid (mostly citric acid) in order to convert them to free fatty acids. In this process mainly the c9,t11 and the t10,c12-CLA isomers are formed in almost equal amounts.
Some improvements of the commercial alkaline isomerisation include, for example, the use of organic solvents with a high boiling point, like ethylene glycol and propylene glycol, instead of water. This way, the reaction can be conducted under the boiling point of the solvent (at 130-150° C. compared to 200-250° C. in the aqueous process), which leads to a better temperature and pressure control, and shorter reaction times (2.5-6 hours). Moreover, the selectivity to the c9,t11- and the t10,c12-CLA isomer is higher (Saebo et al, 2002).
Homogeneous, basic catalysts can also be used for the production of conjugated CLA esters. In this process, alkali metal alcoholate catalysts are used, which in contrast to KOH and NaOH, do not hydrolyze the ester bond. The use of high amounts of bases is often a problem in the industrial process because this lead to corrosion of the reactor.
In the context of the alkaline isomerisation, esterification of conjugated PUFAs/CLAs or either transesterification of conjugated PUFA/CLA alkyl esters is required to obtain triacylglycerols (TAGs) containing conjugated PUFA/CLA.
Conjugated PUFA Production Using Homogeneous Metal Complexes
Different homogeneous catalysts have been tested for the preparation of conjugated fatty acids and oils:                Cr-complexes, such as arene-Cr(CO)3 complexes and Cr(CO)6 (Frankel, 1970),        Rh-complexes, for example RhCl3, [(C6H5)3P]3RhCl (DeJarlais and Gast, 1971ab; Singer et al, 1972, 1977), [(C6H5)3P]2RhCOCl (Singer et al, 1972) and [RhCl(C8H14)2]2 (Larock et al, 2001),        Pt-complexes, as cis-Cl2[(C6H5)3P]2PtSnCl2 and PtCl2(PPh3)2 (DeJarlais and Gast, 1971, Larock et al, 2001), and        Ru-complexes, such as Ru(η6-naphthalene)(η4-cycloocta-1,5-diene) (Pertici et al., 1999) and RuHCl(CO)(PPh3)3 (Larock et al., 2001).        
The substrates can be fatty acids like linoleate as well as polyunsaturated oils, for example soybean or safflower oils. Like the isomerization reaction in strong alkali, several conjugated products are formed. These systems are characterised by low reaction temperatures, high selectivity towards CLA, and the fact that TAGs enriched in CLA can be produced directly. However, the main drawback of these systems is that the catalysts are soluble homogeneous metal complexes, which are not environmentally friendly and difficult to separate from the reaction medium or products. The reuse of such catalysts and the ligands and the use of high amounts of solvents are often problematic. In the particular case when the conjugated PUFA/CLA product will be used in food applications, the choice of the solvent will be limited and also only very low metal contamination levels are acceptable.
Conjugated PUFA Production Via Heterogeneous Catalysis
Heterogeneous catalysis (i.e. a metal catalyst deposited on a porous anorganic or carbon support with a large internal surface) constitutes an attractive strategy for sustainable CLA production, as the catalyst can be separated and reused easily. Although some heterogeneous processes for isomerisation of linoleic acid or methyl linoleate have already been described in literature, low productivity is the main drawback.
Different metals and supports have been screened for the production of CLA, including:                a Ru/C catalyst in the isomerization of methyllinoleate (Mukesh et al, 1985; Narasimhan et al, 1985; Deshpande et al, 1985)        a Rh/C catalyst for the isomerization of methyllinoleate (Deshpande et al, 1985)        Ru on different supports (γ-Al2O3, SiO2Al2O3, C and MgO) and in combination with Ni (Mukesh et al, 1988).        
However, besides isomerization also hydrogenation (formation of oleate, elaïdate and stearate), polymerization and coke formation was observed.
Bernas et al (2003) and Bernas et al (2004) screened Ru, Ni, Pd, Pt, Rh, Ir, Os, and bimetallic Pt—Rh supported by activated carbon, Al2O3, SiO2Al2O3, MCM-22, H-MCM-41, H-Y and H-BETA for the isomerization reaction of linoleic acid to CLA. In order to enhance the isomerization reaction a two-step process was used. In a pre-activation step the catalyst surface is first saturated with hydrogen and then the isomerization reaction of linoleic acid to CLA occurs under a N2 atmosphere. However, significant quantities of hydrogenated products, such as oleic acid, were formed.
Kreich and Claus (2005) described a highly selective method for the synthesis of CLAs over heterogeneous silver catalysts and in the constant presence of hydrogen. Also, the use of heterogeneous gold catalysts were tested in the isomerization of linoleic acid under constant hydrogen flow. Depending on the Au catalyst used, isomerization or hydrogenation is favored (Bauer et al, 2009; Simakova et al., 2010).
One of the main disadvantages of the heterogeneous catalyst based isomerization processes is that the productivities in the heterogeneous processes are very low compared to the industrial process using homogeneous bases. Another difficulty in the heterogeneous catalyzed process is the competition between isomerization and hydrogenation. While isomerization can take place in both directions (i.e. from conjugated to isolated double bonds and vice versa), hydrogenation is a consecutive reaction which only goes in one direction and lowers the CLA yield. On the one hand, hydrogen is needed in order to form the half-hydrogenated intermediates which leads to the isomerization of linoleic acid to CLAs, on the other hand too high levels of hydrogen will lead to the formation of unwanted hydrogenated products. Hence, the direct production of CLA using heterogeneous catalysts is a difficult and complicated process.
In this respect, partially hydrogenated vegetable oils contain higher levels of CLAs, indicating that during the hydrogenation of vegetable oils (using heterogeneous catalysts) CLAs are formed (Mossoba et al., 1991; Banni et al., 1994). By finetuning the hydrogenation process increasing levels of CLAs can be accumulated. The isomerization/hydrogenation ratio can be influenced by the catalyst used as well as the reaction conditions. High CLA accumulation requires conducting the hydrogenation at a high temperature, a low hydrogen pressure, a low agitation rate and a high catalyst level (Jung et al., 2001, 2002). However, these conditions also favor the formation of the unwanted C18:1 trans-isomers, which are known to increase the risk of cardiovascular diseases.
Finally, Chorfa et al (2010) described the hydrogenation/isomerization of safflower oil using a rhodium loaded mesoporous molecular sieve. The reaction was conducted at 180° C. and low hydrogen pressure (0.3 bar). The main isomers formed are the c9,t11-, t10,c12- and t,t-CLAs.
In short, the production of conjugated polyunsaturated fatty acids and derivatives thereof, known in the art, has several disadvantages:                In the alkaline conversion of nonconjugated PUFA, the alkali bases, solvents and acids used are disadvantageous from an ecological and economic point of view. Also an extra processing step is needed to neutralise and/or remove the alkaline catalyst. Furthermore, a mixture of different conjugated PUFA isomers is obtained instead of a single PUFA isomer. Also, the use of conjugated PUFA, such as CLA, in food application requires conjugated PUFA enriched TAGs (and not as free fatty acids or FAMEs), which cannot be obtained directly by the use of homogeneous bases due to the saponification. Thus, an extra time-consuming esterification or transesterification of conjugated PUFA (CLA) (as fatty acid or methylester derivative thereof) is needed.        The main drawback of using homogeneous metal complexes is that these catalysts are soluble in the reaction medium, which makes them difficult to separate and is not environmentally friendly. In addition, when the conjugated PUFA or CLA product will be used in food applications, the choice of the solvent will be limited and very low metal contamination levels are required (from food safety point of view and to minimize the oxidation of the unsaturated fatty acids).        The main disadvantage in the heterogeneous production of conjugated PUFA or derivatives thereof, particularly in the presence of H2 or similar compounds, is the competition between hydrogenation and isomerization. Although changing the process conditions can aid in minimizing the hydrogenation reaction and hence the formation of hydrogenated byproducts, the reaction conditions which have a positive influence on the isomerization reaction also favor the production of harmful C18:1 trans-isomers.        
There hence remains a need for novel methods to produce conjugated polyunsaturated fatty acids and derivatives thereof, such as the alkylesters thereof (e.g. PUFA methylester) or glycerides, particularly triglycerides, comprising said conjugated PUFA.
Accordingly, the present invention provides a new heterogeneous isomerisation catalyst for the synthesis of conjugated PUFA in the absence of hydrogen, as well as novel methods for the synthesis of conjugated PUFA using said heterogeneous catalyst, particularly in the absence of hydrogen.